Muscle Fiber Degeneration Research Articles

NOTCH3 is non-enzymatically fragmented in inherited cerebral small-vessel disease

The small-vessel disorder cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) arises from mutations in the human gene encoding NOTCH3 and results in vascular smooth muscle cell degeneration, stroke, and dementia. However, the structural changes in NOTCH3 involved in CADASIL etiology are unclear. Here, we discovered site-specific fragmentation of NOTCH3 protein in pathologically affected vessels of human CADASIL-affected brains. EM-based experiments to pinpoint NOTCH3 localization in these brains indicated accumulation of NOTCH3 fragmentation products in the basement membrane, collagen fibers, and granular osmiophilic material within the cerebrovasculature. Using antibodies generated against a disease-linked neo-epitope found in degenerating vascular medium of CADASIL brains, we mapped the site of fragmentation to the NOTCH3 N terminus at the peptide bond joining Asp80 and Pro81 Cleavage at this site was predicted to separate the first epidermal growth factor (EGF)-like domain from the remainder of the protein. We found that the cleavage product from this fragmentation event is released into the conditioned medium of cells expressing recombinant NOTCH3 fragments. Mutagenesis of Pro81 abolished the fragmentation, and low pH and reducing conditions enhanced NOTCH3 proteolysis. Furthermore, substitution of multiple cysteine residues of the NOTCH3 N terminus activated proteolytic release of the first EGF-like repeat, suggesting that the elimination of multiple disulfide bonds in NOTCH3 accelerates its fragmentation. These characteristics link the signature molecular genetic alterations present in individuals with CADASIL to a post-translational protein alteration in degenerating brain arteries. The cellular consequences of these pathological NOTCH3 fragments are an important area for future investigation.

South American Rattlesnake (Crotalus durissus spp.) Envenomation in Dogs in the Semiarid Region of Brazil

Background: South American rattlesnake (Crotalus durissus spp.) envenomation is rarely reported in small animals and livestock in Brazil. Minor swelling at the snakebite site, skeletal muscle, and renal damage, and severe neurological signs characterize the crotalic envenomation. This case report aims to present epidemiological, clinical, and pathological data of two cases of Crotalus durissus spp envenomation in dogs in the Northeast of Brazil.Cases: Envenomation by Crotalus durissus spp. was recorded in two dogs in Patos, State of Paraíba, Brazil. In Case 1, the dog presented flaccid paralysis, hyporeflexia, a deficit of cranial nerves, epistaxis, and gingival hemorrhages. Laboratory assay showed proteinuria, myoglobinuria, regenerative thrombocytopenia, and increased serum activities of creatine kinase (CK), aspartate aminotransferase (AST), alanine aminotransferase (ALT), and alkaline phosphatase (ALP). The dog was medicated with crotalic antivenom and wholly recovered from local and systemic clinical signs. In Case 2, the dog died and was detected fang marks at the ventral region of the left mandible (two small parallel perforations spaced 2.0 cm apart) at the snakebite site. Cyanosis of the oral cavity, congestion, and hemorrhages in several organs were observed at necropsy. Tubular nephrosis, muscular necrosis, hepatocytes swelling were observed. The owners witnessed snakebites, and the rattlesnakes (Crotalus durissus spp.) identified by the rattle at the end portion of the tail in both cases. Discussion: Natural South American rattlesnake envenomation presents complex clinical signs that makes diagnosis a challenge for veterinary practitioners. The criteria for the correct diagnosis and observed in the two dogs include witness of the snakebite, identification of the snake, detection of fang marks, clinical-pathological findings, and therapeutic response to treatment with specific anti-venom. The dog’s owners did not identify the subspecies of rattlesnakes; however, Crotalus durissus cascavella and Crotalus durissus collilineatus are the only species found in the Northeast region of Brazil. Crotoxin is the primary toxic component of South American rattlesnake, which induces neuromuscular blockage, and neurological signs (skeletal muscle flaccid paralysis, apathy, hyporeflexia, cranial nerve deficits). These clinical signs are similar to those observed in the two dogs. Respiratory distress, cyanosis, pulmonary edema, and hemorrhage are secondary to respiratory muscle paralysis and also detected in a dog (Case 2) with crotalic envenomation. Minor local swelling at the snakebite site, myotoxicity observed in both dogs (high serum activities of CK and AST - Case 1), degeneration and necrosis of muscle fibers - Case 2), and fang marks observed in Case 2, strengthen the diagnosis of Crotalus durissus envenomation. Nephrotoxicity was also detected in both dogs (increased specific gravity of urine - Case 1 and myoglobin deposition and degeneration of renal epithelial tubular cells - Case 2). Coagulative disorders and hepatotoxicity are infrequently in domestic animals and humans with crotalic envenomation. High serum activities of ALP and ALT in Case 1, and swelling of hepatocytes in Case 2, suggest liver damage associated with the crotalic envenomation. The differential diagnosis of South American rattlesnake envenomation should be included in dogs with acute neuromuscular flaccid paralysis, associated or not with bleeding disorders, myoglobinuria, and acute kidney injury.

Histological features of masticatory muscles after botulinum toxin A injection into the right masseter muscle of dystrophin deficient (mdx-) mice

Objective/backgroundThe most frequently used animal model for human DMD (Duchenne muscular dystrophy) research is the mdx mouse. In both species, characteristic histological changes like inflammation, muscle fiber degeneration and fibrosis are the same, but in contrast to humans, in mdx mice, phases of muscle fiber degeneration are compensated by regeneration processes. AimTherefore, the interest of this study was to evaluate histological features in masticatory muscles after BTX-A injection into the right masseter muscle of wild type and dystrophic (mdx) mice, illustrating de- and regeneration processes induced by this substance. Material and methodsThe right masseter muscle of 100 days old healthy and mdx mice were selectively paralyzed by a single intramuscular BTX-A injection. Masseter as well as temporal muscle of injection and non-injection side were carefully dissected 21 days and 42 days after injection, respectively, and fiber diameter, cell nuclei position, necrosis and collagen content were analyzed histomorphologically in order to evaluate de- and regeneration processes in these muscles. Statistical analysis was performed using SigmaStat Software and Mann Whitney U-test (significance level: p < 0.05). ResultsAt both investigation periods and in both mouse strains fiber diameter was significantly reduced and collagen content was significantly increased in the right injected masseter muscle whereas fiber diameters in mdx mice were much smaller, and these differences were even more apparent at the second investigation period. Necrosis and central located nuclei could generally be found in all mdx mice muscles investigated with an amount of centronucleation exceeding 60%, and a significant increase of necrosis six weeks after injection. In wild type mice central located nuclei could primarily be found in the treated masseter muscle with a portion of 2.7%, and this portion decreased after six weeks, whereas in mdx mice a decrease could also be seen in the non-injected muscles. In contrast, in wild type mice necrosis was not apparent at any time and in all muscles investigated. ConclusionFrom our results it can be concluded that in mdx mice masticatory muscles de- and regeneration processes were extended, triggered by a selective BTX-A injection, or mdx mice at this age, independently of BTX-A treatment, went through another cycle of de- and regeneration as a characteristic of this disease.

Cardioprotective effect of hemin in isoprenaline-induced myocardial infarction: role of ATP-sensitive potassium channel and endothelial nitric oxide synthase.

Ischemic heart disease is a common cardiac health problem. Despite the significant advances in prevention and treatment of this disorder, its incidences and complications are very serious. So, the search for more antioxidants and anti-inflammatory agents with cardioprotective effects is an urgent task. We aimed to evaluate the effects of a heme oxygenase 1 (HO1) inducer, hemin (HEM), on isoprenaline (ISO)-induced myocardial damage. Forty-five Wistar albino rats were used. Animals were treated with HEM (25mg/kg/day) i.p. for 5days and injected with ISO (150mg/kg/day) i.p. on 4th and 5th day of the experiment. Detection of the role of ATP-sensitive potassium channel (KATP ) was performed by administration of glibenclamide (GP) (5mg/kg/day) orally 2h before HEM. Moreover, the role of endothelial nitric oxide synthase (eNOS) was detected by coadministration of Nitro- ω-L-arginine (L-NNA) (25mg/kg/day) for 5days. The ISO group showed increase in heart weight, cardiac enzymes, tumor necrosis factor alpha (TNFα), and malondialdehyde (MDA) with decrease in reduced glutathione (GSH), HO1, and total antioxidant capacity (TAC). In addition, there were increases in Bcl-2 associated X protein (Bax) and cleaved caspase-3, but decreases in B-cell lymphoma-2 (Bcl-2) and eNOS. Moreover, the histopathological examination ofthe ISO group showed degeneration of the cardiac muscle fibers and marked infiltration of the inflammatory cells. The biochemical and histopathological changes induced by ISO were markedly ameliorated in the HEM plus ISO group. This protective effect was diminished with coadministration of GP or L-NNA; thus, KATP and eNOS might mediate HEM cardioprotection.

Scuticociliatosis caused by Uronema sp. in ten different ornamental aquarium reef fish in Brazil.

Scuticociliatosis, caused by an opportunistic ciliate protozoan, is responsible for significant economic losses in marine ornamental fish. This study reports the occurrence of Uronema spp., parasitizing ten species of marine reef fish at an ornamental fish wholesaler: Blue green damselfish (Chromis viridis), Vanderbilt's Chromis (Chromis vanderbilti), Pennant coralfish (Heniochus acuminatus), Threespot angelfish (Apolemichthys trimaculatus), Goldspotted angelfish (Apolemichthys xanthopunctatus), Sea goldie (Pseudanthias squamipinnis), Orchid dottyback (Pseudochromis fridmani), Threadfin butterflyfish (Chaetodon auriga), Vagabond butterflyfish (Chaetodon vagabundus), and Bluecheek butterflyfish (Chaetodon semilarvatus). Diseased fish showed disorders such as hemorrhages and ulcerative lesions on the body surface. Histopathological analysis of the muscle, liver, gut, kidney, spleen, gills, and stomach revealed hemorrhages and degeneration of muscle fiber, vacuolar degeneration of hepatocyte, inflammatory process and granuloma in the liver, atrophy of intestinal villi, inflammatory process and granuloma in the kidney, melanomacrophage centers, as well as inflammatory process in the spleen, epithelial cells hyperplasia and granuloma formation in the gills, and vacuolar degeneration and eosinophils in the stomach. Due to the severity of the disease, it is necessary to implement biosecurity measures with rapid and accurate diagnosis to minimize the risk of economic losses caused by Uronema spp.

Isolation, Culture, and Differentiation of Primary Myoblasts Derived from Muscle Satellite Cells

The skeletal muscle is key for body mobility and motor performance, but aging and diseases often lead to progressive loss of muscle mass due to wasting or degeneration of muscle cells. Muscle satellite cells (MuSCs) represent a population of tissue stem cells residing in the skeletal muscles and are responsible for homeostatic maintenance and regeneration of skeletal muscles. Growth, injury, and degenerative signals activate MuSCs, which then proliferate (proliferating MuSCs are called myoblasts), differentiate and fuse with existing multinuclear muscle cells (myofibers) to mediate muscle growth and repair. Here, we describe a protocol for isolating MuSCs from skeletal muscles of mice for in vitro analysis. In addition, we provide a detailed protocol on how to culture and differentiate primary myoblasts into myotubes and an immunofluorescent staining procedure to characterize the cells. These methods are essential for modeling regenerative myogenesis in vitro to understand the dynamics, function and molecular regulation of MuSCs.

Oesophageal sarcocystosis in an Indian female buffalo (Bubalus bubalis) - A case study

An adult female Murrah water buffalo (Bubalus bubalis) carcass from Cattle and Buffalo Farm, ICAR-Indian Veterinary Research Institute (ICAR-IVRI), Izatnagar, Bareilly, Uttar Pradesh, India (28°10'N and 78°58'E) was sent to Postmortem Examination Facility of Division of Pathology, ICAR-IVRI and necropsy was carried out. On systematic examination, large, opaque white, boiled rice-like cysts and typical fusiform shape with wide middle and tapered ends, and embedded in the muscularis externa, underneath the serosa and visible through the serosal surface of the oesophagus. They were easily separated from oesophagus. Sarcocysts were noticed in entire oesophagus from pharyngeal part to abdominal oesophagus. The cysts were of different size (ranges from 7-20 mm length and 2-5 mm width) with uniform shape. The contents of cyst showed crescent shaped bradyzoites and few metrocytes. On histopathological examination, cyst wall was thin and smooth with 1-3 pm thickness. Ground substance beneath the cyst wall contained metrocytes. Inner compartmentalization contained various intensities of bradyzoites. Hyaline degeneration and atrophy of oesophageal muscle fibers were noticed. In the present study, based on the distinctive shape (fusiform shaped with wide middle and tapered ends) and size (7-20 mm in length and 2-5 mm in width) of cysts and microscopic cyst wall thickness (thin and smooth cyst wall with 1-3 pm width), the sarcocyst was confirmed as macrosarcocyst of S. fusiformis.

Effects of Carbachol on the Excitatory Muscarinic Cholinergic Receptors in Urinary Bladder and Plasma Inflammatory Mediators in Anesthetized Rats

To observe the effects of muscarinic cholinergic receptor agonist carbachol on urinary bladder function and plasma inflammatory mediators in anesthetized rats. A total of 20 rats were randomly divided into carbachol group and solvent control group, 10 rats in each group. The detection of bladder wall thickness and histological observation were conducted by light microscopy, the changes of urinary bladder function and plasma inflammatory mediators in rats were compared between the two groups. The maximum bladder capacity, static bladder pressure, compliance, bladder wet weight and bladder wall thickness in the carbachol group were significantly higher than those in the solvent control group. Light microscopic evaluation revealed that the detrusor cells in the solvent control group appeared to be in order in terms of size, arrangement and shape, while in the carbachol group the detrusor cells exhibited hypertrophy together with disordered arrangement, diverse morphology and vacuolar degeneration of muscle fiber. The levels of TNF-α and IL-10 in plasma of the carbachol group were significantly lower than those of the solvent control group. In anesthetized rats, the stimulation of muscarinic cholinergic receptors can significantly reduce urinary frequency, increase the volume of bladder storage and decrease systemic inflammatory response.

DESMOID-TYPE FIBROMATOSIS OF THE BUCCAL MUCOSA: A CASE REPORT

Desmoid-type fibromatosis is a rare benign mesenchymal lesion that originates from musculoaponeurotic structures and rarely occurs in the head and neck region. The 32-year-old male patient was referred for evaluation of a submucosal lesion located in the left buccal mucosa, which was hardened, was painless, and had 3 months of evolution. Ultrasonography showed a well-defined, anechoic, and hypovascular mass. The histopathologic analysis showed proliferation of elongated, spindle-shaped cells of uniform appearance exhibiting condensed chromatin and eosinophilic cytoplasm with imprecise limits, sometimes with fasciculate pattern, associated with areas of prominent fibrosis with thick bundles of collagen. The proliferation produces dissociation and degeneration of muscle fibers. Immunohistochemical reaction revealed, in tumor cells, negativity for protein S100 and CD34 besides immunoexpression focal positive for α actin and β catenin. The cells strongly expressed vimentin. Morphologic and immunohistochemical findings were compatible with desmoid type fibromatosis. The patient is under clinical follow-up of 8 months without relapse. Desmoid-type fibromatosis is a rare benign mesenchymal lesion that originates from musculoaponeurotic structures and rarely occurs in the head and neck region. The 32-year-old male patient was referred for evaluation of a submucosal lesion located in the left buccal mucosa, which was hardened, was painless, and had 3 months of evolution. Ultrasonography showed a well-defined, anechoic, and hypovascular mass. The histopathologic analysis showed proliferation of elongated, spindle-shaped cells of uniform appearance exhibiting condensed chromatin and eosinophilic cytoplasm with imprecise limits, sometimes with fasciculate pattern, associated with areas of prominent fibrosis with thick bundles of collagen. The proliferation produces dissociation and degeneration of muscle fibers. Immunohistochemical reaction revealed, in tumor cells, negativity for protein S100 and CD34 besides immunoexpression focal positive for α actin and β catenin. The cells strongly expressed vimentin. Morphologic and immunohistochemical findings were compatible with desmoid type fibromatosis. The patient is under clinical follow-up of 8 months without relapse.

Effect of feed restriction timing on live performance, breast myopathy occurrence, and muscle fiber degeneration in 2 broiler chicken genetic lines

During recent years, research on meat quality in poultry has aimed to evaluate the presence and consequences of breast myopathies as well as the factors which can affect their occurrence by modifying the growth rate. A total of 900 broiler chickens were reared until slaughter (48 D) to evaluate the effect of 2 genetic lines (A vs. B) and feeding plans (ad libitum [AL], early restricted [ER], from 13 to 23 D of age, and late restricted [LR], from 27 to 37 D of age; restriction rate: 80%) on performance, meat quality, and breast muscle myopathies. Calsequestrin and vascular endothelial growth factor (VEGF) expressions, and muscle fiber degeneration (MFD) were recorded at 22, 36, and 48 D. Chickens in the AL treatment had greater final live (P < 0.01) and carcass weights and proportion of pectoralis major muscle (P = 0.04) compared to chickens in the LR treatment, whereas chickens in the ER treatment had intermediate final live (3,454g) and carcass weights, and proportion of pectoralis major muscle (25.6%). Chickens of line A were heavier than chickens of line B (P < 0.001), and had a greater feed conversion rate. Chickens of line A also had a greater dressing out percentage (P < 0.001), but a lower proportion of pectoralis major muscle (P = 0.04), as well as a greater meat pH (P < 0.001), meat cooking losses (P < 0.01), and shear force of the pectoralis major muscle (P = 0.03). Calsequestrin and VEGF mRNA were significantly lower in ER and LR chickens compared to AL chickens after feed restriction and during refeeding (P < 0.05). MFD scores increased with chicken age (P < 0.001) and differed between genetic lines (P < 0.001). Neither feeding plan nor genetic line affected the occurrence of white striping or wooden breast condition.

Mineralocorticoid Receptor Signaling Contributes to Normal Muscle Repair After Acute Injury.

Acute skeletal muscle injury is followed by a temporal response of immune cells, fibroblasts, and muscle progenitor cells within the muscle microenvironment to restore function. These same cell types are repeatedly activated in muscular dystrophy from chronic muscle injury, but eventually, the regenerative portion of the cycle is disrupted and fibrosis replaces degenerated muscle fibers. Mineralocorticoid receptor (MR) antagonist drugs have been demonstrated to increase skeletal muscle function, decrease fibrosis, and directly improve membrane integrity in muscular dystrophy mice, and therefore are being tested clinically. Conditional knockout of MR from muscle fibers in muscular dystrophy mice also improves skeletal muscle function and decreases fibrosis. The mechanism of efficacy likely results from blocking MR signaling by its endogenous agonist aldosterone, being produced at high local levels in regions of muscle damage by infiltrating myeloid cells. Since chronic and acute injuries share the same cellular processes to regenerate muscle, and MR antagonists are clinically used for a wide variety of conditions, it is crucial to define the role of MR signaling in normal muscle repair after injury. In this study, we performed acute injuries using barium chloride injections into tibialis anterior muscles both in myofiber MR conditional knockout mice on a wild-type background (MRcko) and in MR antagonist-treated wild-type mice. Steps of the muscle regeneration response were analyzed at 1, 4, 7, or 14 days after injury. Presence of the aldosterone synthase enzyme was also assessed during the injury repair process. We show for the first time aldosterone synthase localization in infiltrating immune cells of normal skeletal muscle after acute injury. MRcko mice had an increased muscle area infiltrated by aldosterone synthase positive myeloid cells compared to control injured animals. Both MRcko and MR antagonist treatment stabilized damaged myofibers and increased collagen infiltration or compaction at 4 days post-injury. MR antagonist treatment also led to reduced myofiber size at 7 and 14 days post-injury. These data support that MR signaling contributes to the normal muscle repair process following acute injury. MR antagonist treatment delays muscle fiber growth, so temporary discontinuation of these drugs after a severe muscle injury could be considered.

Assessment of DNA integrity through MN bioassay of erythrocytes and histopathological changes in Wallago attu and Cirrhinus mirigala in response to freshwater pollution.

The aim of this study was to determine the level of contamination and genotoxic impact through micronucleus assay and histopathology in Wallago attu and Cirrhinus mrigala procured from the polluted site of the River Chenab at industrial and sewage waste disposal. The water sample was found viciously contaminated with heavy metals i.e. Ni, Cr, Mn, Co, Pb, Hg, Zn, Sn, Cu while all other physio-chemical variables crossed the suggested limits of WHO. The heavy metals load induced histopathological alterations were correlated to environmental degradation and the productivity of this biological system. W. attu and C. mrigala harvested from contaminated sites of the river indicated higher intensity of DNA damage through micronucleus induction and nuclear abnormalities with 5.46 ± 0.17, 1.23 ± 0.08 and 4.2 ± 0.11, 0.4 ± 0.04‰ respectively. Muscle sections of W. attu and C. mrigala harvested from the polluted section of river demonstrated the necrosis, degeneration of muscle fibers, intra-fibular edema and release of the blood into the tissues due to the bursting of blocked of the blood vessels. Dermal layers showed degeneration of the collagen bundles those were found loose or collapsed in some regions. Photomicrography also revealed vacuolar degeneration in muscle tissues and atrophy of muscle bundles. Intra fibular edema and splitting of muscle fibers were also seen along with bioaccumulation of toxicants. W. attu showed maximum incidence of alterations with highest histopathological alteration index related to environmental degradation. Control fish samples showed normal muscle tissues with normal equally spaced muscle bundles and myotomes.

Histological and Immunohistochemical Study on the Possible Effect of Mangosteen and Mesenchymal Stem Cells on Isoproterenol Induced Myocardial Infarction in Adult Male Albino Rats

Background: Myocardial infarction (MI) is one of the causes of cardiovascular diseases that lead to death despite of the great advancements in the medical interventions.Aim of the work: To assess the possible effect of mangosteen and mesenchymal stem cells (MSCs) in Isoproterenol (ISO)-induced MI in adult male albino rats, monitored by histological and immunohistochemical methods.Materials and Methods: Fifty-five adult male albino rats were divided into: control group (GI), group II (ISO), group III (Mangosteen+ISO), group IV (ISO+MSCs), group V (Mangosteen+Isoproterenol+MSCs). ISO was given subcutaneously, twice (250mg/kg/d), mangosteen was given orally (18mg/200gm) and MSCs (5x106 cells in 1mL of PBS) were injected intracardiac. Myocardial sections were stained with H&E, Masson’s trichrome and immunohistochemical stain for caspase-3 and vascular endothelial growth factor (VEGF). The mean area % of collagen fibers and immunoreactivity of caspase-3 and VEGF were measured by the image analyzer. The statistical analysis was applied by using ANOVA.Results: Myocardial sections of G II revealed discontinuity and degeneration of cardiac muscle fibers, pyknotic nuclei, inflammatory cellular infiltration and congested blood vessels. There were increased collagen deposition and significant increase in caspase-3 immunoreactivity. Mangosteen administration (G III) showed partial improvement in cardiac muscle fibers and reduced inflammatory cellular infiltration.MSCs treatment (G IV) resulted in obvious decline in myocardial damage, apoptosis with significant reduction of collagen deposition and significant increase in VEGF immunoreactivity. Better results are obtained when MSCs and mangosteen were combined (G V).Conclusion: Mangosteen exerts a synergistic effect with MSCs against ISO-induced MI.

From the Literature

From the Literature

Multiorgan histopathological changes in the juvenile seabream Sparus aurata as a biomarker for zinc oxide particles toxicity.

Zinc oxide nanoparticles are widely used in some domains (cosmetics, pharmaceuticals optical devices, and agricultural field) due to their physical, optical, and antimicrobial properties. However, the release of ZnO-NPs into the environment may affect organisms like fish with potential consequences for human health. Histological approaches of the acute effects of these materials on fish are scarce; thus, the present study aimed to investigate the potential toxic effects of acute exposure to ZnO particles in marine environments, by assessing histological changes in the gills, liver, spleen, and muscle of gilthead seabream (Sparus aurata) juveniles. Thus, fish were exposed for 96 h, via water, to 1 mg L-1 of ionic zinc and zinc oxide particles (1.1, 1.2, and 1.4 μm of size). Histological examination revealed gills as the most affected organ, followed by liver, muscle, and spleen. In the gills, histopathological changes included hyperplasia of epithelial cells, fusion of the secondary lamellae, and lifting of the lamellar epithelium with edema. In the liver, lipid vacuolation of several degrees, necrosis of hepatic and pancreatic tissues, blood congestion in sinusoids and hepatoportal vessels, presence of cellular infiltrate, and melano-macrophages diffusion was found. Muscle showed degeneration, atrophy, thickening and necrosis of muscle fibers with edema between them, and presence of melano-macrophages in the muscle layer. Spleen was the less damaged organ, displaying congested blood, white pulp increase/rupture, and bigger and darker melano-macrophage aggregates in the splenic stroma. These results underline that the size of particles plays a determinant role in their potential pernicious effects. A short-term exposure caused major histopathological changes in relevant organs of S. aurata juveniles, possibly affecting their function.

Metabolic and morphological disorientations in the liver and skeletal muscle of mice exposed to hexavalent chromium

Chromium exposure (10 mg/kg b.w/day for 30 days) produced hypoglycaemia in Swiss albino mice associated with increased glycogenolytic activity of hepatic and muscular tissue and suppressed glycolytic activity of the liver. Lactate dehydrogenase activity was decreased in hepatic tissue following exposure to chromium. Chromium significantly inhibited the activities of succinate dehydrogenase and NADH dehydrogenase in all the studied tissues. Moreover, suppressed malate dehydrogenase activity in chromium stressed muscle indicates less energy supply to that tissue, whereas its increased activity may aid substrates for gluconeogenesis in the liver. Chromium caused marked alteration in the liver and muscle protein contents and increased free amino acid nitrogen level. Proteolytic enzyme activities like trypsin, cathepsin and pronase were also significantly altered. Additionally, the transaminase enzyme activities were elevated in all the tissues after chromium exposure. A compensatory mechanism was initiated by inducing fatty acid synthase activity in hepatic tissue. The lipogenesis was enhanced in the liver which might involve the isocitrate dehydrogenase activity as well as overproduction of cholesterol and triglycerides. In response to these, the reverse transport of cholesterol from the blood to the liver was hampered due to less production of HDL cholesterol in chromium toxicity. Histopathological changes in the liver showed steatosis along with alteration in radiating pattern of hepatic cells in chromium treated mice. Additionally, skeletal muscle fibre degeneration was also evident. Over accumulation of chromium was noted in the studied tissues after chromium treatment at the present dose and duration. The study suggests potential threatening effect of hexavalent chromium on the liver and skeletal muscle metabolism.

Association between TDP-43 and mitochondria in inclusion body myositis

Inclusion body myositis (IBM) is the most common cause of primary myopathy in individuals aged 50 years and over, and is pathologically characterized by protein aggregates of p62 and mislocalized cytoplasmic TDP-43, as well as mitochondrial abnormalities in affected muscle fibers. Our recent studies have shown the accumulation of TDP-43 in mitochondria in neurons from patients with amyotrophic lateral sclerosis (ALS) and frontotemporal degeneration (FTD), and revealed mitochondria as critical mediators of TDP-43 neurotoxicity. In this study, we investigated the association between mitochondria and TDP-43 in biopsied skeletal muscle samples from IBM patients. We found that IBM pathological markers TDP-43, phosphorylated TDP-43, and p62 all coexisted with intensively stained key subunits of mitochondrial oxidative phosphorylation complexes I–V in the same skeletal muscle fibers of patients with IBM. Further immunoblot analysis showed increased levels of TDP-43, truncated TDP-43, phosphorylated TDP-43, and p62, but decreased levels of key subunits of mitochondrial oxidative phosphorylation complexes I and III in IBM patients compared to aged matched control subjects. This is the first demonstration of the close association of TDP-43 accumulation with mitochondria in degenerating muscle fibers in IBM and this association may contribute to the development of mitochondrial dysfunction and pathological protein aggregates.

Effect of electroacupuncture at "Weizhong" (BL40) on expression of collagen I and matrix meta-lloproteinases 2 in rats with lumbar multifidus muscle injury

To observe the effect of electroacupuncture (EA) at "Weizhong" (BL40) on histopathological changes and expression of extracellular matrix (ECM) component Collagen Ⅰ， matrix metalloproteinases 2 (MMP2), MyoD and Pax7 proteins of lumbar muscle tissues in rats with lumbar multifidus muscle injury (LMMI), so as to explore its underlying mechanisms in improving muscular injury. A total of 24 male SD rats were equally randomized into blank control, model and EA groups. The LMMI model was established by injection of 0.5% Bupivacaine (100 µL/ point) into bilateral multifidus muscles of lumbar 4 and 5 (4 points). EA (2 Hz/100 Hz in frequency, 1－2 mA) was applied to BL40 for 20 min, once a day for 3 days. The morphological changes of the left lumbar multifidus muscle were observed under microscope after H．E. and Masson staining, and the expression of Collagen Ⅰ， MMP2, MyoD and Pax7 of the right lumbar multifidus muscle was determined by Western blot. H．E. staining showed large areas of degeneration and necrosis of muscle fibers, and vacuolar structure formed by degradation of muscle fibers in the model group, and newborn juvenile muscle fibers with different diameters in the EA group. Masson staining showed a large number of morphologically damaged muscle fibers and blue stained collagen fibers in the model group, and significantly reduced collagen fibers as well as new muscle fibers with uneven diameters in the EA group. The expression levels of Collagen Ⅰ， MMP2 and MyoD proteins were significantly up-regulated (P<0.01, P<0.05), and that of Pax7 was considerably down-regulated in the model group relative to the control group (P<0.01). After EA intervention, the expression levels of Collagen Ⅰ was significantly down-regulated (P<0.01), and those of MMP2, MyoD and Pax7 proteins were obviously or further obviously up-regulated in the EA group compared with the model group (P<0.01, P<0.05). EA at BL40 can reduce the degree of skeletal muscle fibrosis to promote the regeneration of the injured multifidus at the early phase, which may be related to its effect in regulating the expression of Collagen Ⅰ and MMP2 proteins.

Influence of Heat Stress on Monocyte Chemoattractant Protein‐1 Expression by Satellite Cells after Rat Skeletal Muscle Injury

Heat stress could promote skeletal muscle regeneration via early recruitment and invasion of macrophages into the necrotic muscle fibers (e.g., Takeuchi et al., 2014; Hatade et al., 2014). Macrophages play important roles in myoblasts proliferation, differentiation and fusion by producing some cytokines and be involved in muscle regeneration (e.g., Tidball, 2005; Arnold et al., 2007). But in the regeneration process, the mechanisms for the macrophages recruitment via the heat stress is unclear. Monocyte Chemoattractant Protein‐1 (MCP‐1) is involved in the migration, and this chemokine is produced by the satellite cells (e.g., Chazaud et al., 2003). Spatial distributions and chronological changes of the MCP‐1, Pax7 (a specific marker of satellite cells) and CD68 (a pan‐macrophages marker) were examined by using the immunohistochemistry via heat stress after crush injury to the extensor digitorum longus muscle of rats. In the present study, 8‐week old male Wistar rats weighing 180–200g were used. According to the crush method described by Furuta (2001) and our earlier reports (Takeuchi et al., 2014; Hatade et al., 2014), animals were anesthetized by an isoflurane and exposed the muscle. Next, the middle part of the muscle belly was crushed for 30s using forceps, to which a weight (500g) was attached. After the muscle crush injury, animals were randomly divided into heat and non‐heat groups. In the heat group, packs filled with hot water (42 degrees C) were percutaneously applied to the muscle for 20 min soon after the injury and in the non‐heat group were not. At 6, 12 and 24 h (n = 1, respectively) after the injury, animals were sacrificed, muscles were harvested and frozen cross sections (10 μm) of the injured muscle were prepared by cutting with a cryostat. Some sections were used for hematoxylin and eosin staining, the others were used for immunohistochemistry. At 6 h after the injury, macrophages were recruited around the degenerating muscle fibers in the non‐heat group. In the heat group, recruitment of the macrophages was higher than the non‐heat group. Similarly, recruitment and infiltration of macrophages in the heat group were higher than those in the non‐heat group at 12 h and 24 h after the injury. In addition, at 6 h after the injury of the heat group, the MCP‐1 was expressed in the satellite cells, whereas in the non‐heat group it was seen after 12 h, suggesting that the heat stress soon after the skeletal muscle injury might promote more rapid MCP‐1 expression of the satellite cells.Support or Funding InformationNoneThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Histopathological Changes and Antioxidant Enzymes Status in Oxidative Stress Induction Using Sodium arsenite in Rats

Introduction: Arsenic toxicity has posed troublesome health concerns in the world and many of the toxic effects of arsenic are related to its effect on oxidative stress. The aim of the present study is to evaluate histopathological and antioxidant enzymes changes in oxidative stress status induced by sodium arsenite in rats. Materials and Methods: All experiments were carried out in male Wistar rats. Animal were divided into 2 groups of eight animals in each: Rats consumed distilled water (control group). Group 2: Rats consumed a solution of sodium arsenite (100 ppm) daily (arsenic group). At the end of day 28 arsenic exposure, the enzyme level in rat liver and kidney tissues was measured using the assay kits. The remaining liver, kidney, and heart tissue were fixed in 10% neutral-buffered formalin and used for histological observation. Results: The results showed a significant decrease in values of glutathione peroxidase (GPx), superoxide dismutase (SOD), total antioxidant capacity (TAC) and catalase (CAT) in serum and tissue in kidney and liver rat (P < 0.05). But, malondialdehyde (MDA) levels were increased significantly (P < 0.05). Arsenic caused severe degenerative changes in tubular cells and acute tubular necrosis, hepatocyte cell degeneration, severe hemorrhage, and infiltration and formation of Kupffer cells nodules, fragmentation, and degeneration of muscle fibers with pyknotic nuclei in heart tissue. Conclusions: The finding of the present study revealed that the administration of Sodium arsenite caused significant oxidative stress, decreased antioxidant enzymes activity and severe tissue damage.